1. Field of the Invention
The present invention relates to a method for the recovery of .alpha.-L-aspartyl-L-phenylalanine methyl ester, L-phenylalanine and L-aspartic acid from dilute solutions containing .alpha.-L-aspartyl-L-phenylalanine methyl ester (hereafter abbreviated as .alpha.-APM) which comprises recovering .alpha.-APM from a mother liquor from which .alpha.-APM has already been crystallized out of, and also recovering L-phenylalanine and L-aspartic acid after hydrolysis.
More particularly, the present invention relates to a method for recovery of .alpha.-L-aspartyl-L-phenylalanine methyl ester, L-phenylalanine and L-aspartic acid which comprises concentrating part or all of the mother liquor obtained by the solid-liquid separation of a suspension of .alpha.-L-aspartyl-L-phenylalanine methyl ester crystals, either taking a part of the resulting concentrate (Solution A) and adding a mineral acid and methanol thereto or mixing Solution A with a solvent comprising mineral acid, methanol, water, and, optionally, .alpha.-L-aspartyl-L-phenylalanine methyl ester, to recover .alpha.-L-aspartyl-L-phenylalanine methyl ester and/or its acid addition salt analogue (.alpha.-L-aspartyl-L-phenylalanine methyl ester acid salt), and hydrolyzing the remaining concentrate and, if desired, the remaining mother liquor with a mineral acid to recover L-phenylalanine or L-phenylalanine and L-aspartic acid.
2. Discussion of the Background
.alpha.-APM is a peptide sweetener which exhibits a sweetness about 200 times that of sucrose. Because of its sweetness, extremely high flavor quality and low calorie content, .alpha.-APM has been widely used as a dietetic sweetener in recent years. It is expected that world .alpha.-APM demand will exceed 10,000 tons/year by 1995.
.alpha.-APM has been prepared on an industrial scale according to the following methods: (1) condensing an N-protected aspartic anhydride with L-phenylalanine methyl ester in an organic solvent and removing the protecting group in a conventional manner (U.S. Pat. No. 3,786,039); (2) converting .alpha.-L-aspartyl-L-phenylalanine in a mixture of water, methanol and hydrochloric acid into the methyl ester to form .alpha.-APM hydrochloride, followed by neutralizing the hydrochloride to give .alpha.-APM (Japanese Patent Application Laid-Open No. 53-82752); and (3) condensing an N-protected aspartic acid with phenylalanine methyl ester in the presence of an enzyme followed by removal of the protecting group (Japanese Patent Application Laid-Open No. 55-135595).
In the chemical synthesis described in (1) above, impurities including .beta.-isomers (i.e., .beta.-L-aspartyl-L-phenylalanine methyl ester) are necessarily produced as unwanted by-products. In order to selectively remove these impurities, a method for purification is known which comprises contacting impure .alpha.-APM with a hydrohalic acid and performing solid-liquid separation to isolate .alpha.-APM as the hydrohalide salt (4).
The most popular method of industrial preparation of .alpha.-APM is via the methyl ester hydrochloride obtained after esterification, as shown in (2) above. Further, and as shown in (4) above, the purification of .alpha.-APM is often pursued through the hydrohalide. In order to obtain .alpha.-APM from its hydrohalide, including the hydrochloride, it is conventional to dissolve or suspend the hydrohalide of .alpha.-APM in an aqueous medium followed by the addition of aqueous solutions of sodium carbonate, sodium oxide, sodium hydrogencarbonate or ammonia to carry out the neutralization.
In preparing .alpha.-APM on an industrial scale, the mother liquors formed during the course of isolating and purifying .alpha.-APM contain, in addition to un-isolated .alpha.-APM, compounds associated with its production such as .alpha.-L-aspartyl-L-phenylalanine (.alpha.-AP), .beta.-L-aspartyl-L-phenylalanine methyl ester (.beta.-APM,), .beta.-L-aspartyl-L-phenylalanine (.beta.-AP), 5-benzyl-3,6-dioxo-2-piperazineacetic acid (DKP), .alpha.-L-aspartyl(.beta.-methyl)-L-phenylalanine methyl ester (.alpha.-A(M)PM), .alpha.-L-aspartyl(.beta.-methyl)-L-phenylalanine (.alpha.-A(M)P), N-formyl-.alpha.-L-aspartyl-L-phenylalanine methyl ester, and large quantities of inorganic salts due to prior neutralization(s). The type and amount of inorganic salts vary depending upon an alkali used to neutralize the .alpha.-APM hydrohalide such as .alpha.-APM hydrochloride, etc. Generally, the inorganic salts are NaCl, NH.sub.4 Cl, KCl, etc.
The recovery of valuable starting materials, particularly the amino acids used as the main raw materials in the production of .alpha.-APM, from the mother liquors mentioned above greatly contributes to an overall reduction in the production costs of .alpha.-APM. In order to enhance productivity on an industrial scale, it is extremely important to recover these substances. Furthermore, it is advantageous that any recovery process also lead to a reduction in the organic materials present in any waste liquid(s) so a to reduce the overall cost of waste treatment.
The following techniques are known for recovery of .alpha.-APM starting materials: (1) hydrolyzing .beta.-APM with an aqueous mineral acid solution and precipitating L-phenylalanine at Ph of 4 to 8, and then precipitating L-aspartic acid at a Ph of 3 to 1 (Japanese Patent Application Laid-Open No. 48-97812), (2) concentrating one or more filtrates, etc. used in the preparation of .alpha.-APM, hydrolyzing the concentrate, rendering the concentrate acidic to precipitate the salt of L-phenylalanine, and adjusting Ph of the mother liquor to obtain L-aspartic acid (Japanese Patent Application Laid-open No. 57-130958), and (3) concentrating the mother liquor formed during the course of preparing .alpha.-APM below 70.degree. C., hydrolyzing the concentrate to recover L-phenylalanine and L-aspartic acid (Japanese Patent Application Laid-Open No. 63-159355).